Source control and antibiotics are the current mainstay of therapy for intra-abdominal sepsis. However, mortality and morbidity, in the form of persistent critical illness with organ failure, remain high because many patients present well into their clinical course and with advanced sepsis. We therefore still need a greater understanding of the mechanisms leading to impaired bacterial clearance and excessive systemic inflammation during abdominal sepsis. Improved mechanistic understanding of abdominal sepsis is critical to identifying novel targets that have broad therapeutic time windows, with the ideal therapy being one that could be implemented upon presentation to modify the host response and improve outcomes. Lipopolysaccharide (LPS) from Gram-negative bacteria is a well-established driver of host responses to intra-abdominal infection. An intracellular, cytosolic LPS sensing pathway involving caspase-11 has recently been identified to also play a role in host responses to Gram negative infection. Because caspase-11 upregulation and activation occurs in a delayed fashion it has gained considerable interest as a potential therapeutic target. However, the interplay between cell surface LPS sensing by TLR4 and cytoplasmic LPS sensing by caspase-11 is not understood and the critical cell-specific TLR4/caspase-11 functions involved in the host response to polymicrobial intra-abdominal infection are not known. We hypothesize that the interplay between TLR4 and caspase-11 (caspase-4 in humans) is a key determinant of bacterial clearance and systemic inflammation during polymicrobial intra-abdominal sepsis, with LPS sensing coordinated jointly by the TLR4 receptor complex and caspase-11. TLR stimulation increases cytosolic caspase-11 expression, and caspase-11 can be activated by internalized LPS. Our preliminary findings predict that both LPS sensing and downstream functions of TLR4 and caspase-11 are also cell specific after polymicrobial abdominal sepsis. In myeloid cells, such as macrophages there is enhanced bacterial phagocytosis, more efficient bacterial clearance, and increased caspase-11-mediated pyroptosis with passive release of HMGB1 to fuel systemic inflammation. In hepatocytes, the primary cell involved in LPS-uptake and clearance after CLP, TLR4-dependent LPS uptake leads to upregulation and activation of caspase- 11 and active release of HMGB1 in exosomes, without increased pyropotosis. Our preliminary data indicate that hepatocytes are the dominant source of circulating HMGB1 in CLP sepsis, although we propose that excessive leukocyte pyroptosis can impair bacterial clearance while also promoting prolonged inflammation through passive HMGB1 release. In this proposal we will define the cell specific contributions of TLR4 and caspase-11 during cecal ligation and puncture, a clinically relevant model of surgical abdominal sepsis (Aim I). We will focus more detailed mechanistic studies on the unique roles of caspase-11 in driving HMGB1 out of hepatocytes in a novel and active process distinct from pyroptosis (Aim II).